Fabrication of electrodes with high spatial resolution and patterning are now mainstream methods in microfabrication and biosensing. The spatial patterning of different biorecognition molecules (e.g. antibodies, enzymes, aptamers) has not kept pace in practice. By far, the most common methods of biomolecule functionalization of electrodes for routine fabrication of biosensors for physiology are dip-coating, drop-coating, or spin-coating, none of which can be used to functionalize two nearby electrodes (micron scale spacing) with two different biomolecules for multi-analyte sensing.
Top-down approaches such as dip pen lithography or contact printing can be used, but these are hindered by speed, requirement for highly specialized equipment or the need for master templates. Alignment between the electrode and the depositing pen or mask is a primary challenge that becomes more difficult as spatial features decrease in size. Bottom up approaches typically rely on the chemical selectivity of the coupling chemistry to achieve spatially controlled deposition of specific biomolecules, limiting the number of unique sensors that can be created in a small space. In addition, specific coupling chemistries may be limited to the type of electrode material (e.g. thiol linkages to gold) or biomolecule of interest. In order to decorate nearby microelectrodes with different recognition biomolecules, a method needs to be developed to selectively control molecular deposition to each electrode.